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a bright enough signal; currently, the most far red-shifted derivatives of the GFP family emit a maximum of 670nm fluorescence and excite at around 610nm. ‘Our erythrocytes contain haemoglobin, which absorbs light below 650nm,’ said Professor Vladislav Verkhusha from the Albert Einstein College of Medicine. ‘To excite and detect fluorescence efficiently in biological tissue we need fluorescence proteins with both excitation and emission spectra above 650nm.’ So, although the maximum emission value is 670nm, the excitation is still only 610nm, which isn’t ideal for imaging
These imaging
techniques will become more popular as biologists adopt these new probes
tissue. ‘They work partially but not perfectly,’ Verkhusha noted. ‘Fluorescence that emits below 650nm will not be visible, because it will be absorbed by haemoglobin in the tissue.
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There have been many research groups over the last two decades that have tried to improve the excitation and emission in the near- infrared (NIR), but this is the maximum that is achievable now.’ Professor Verkhusha and his team at the Albert Einstein College of Medicine have developed a new group of NIR fluorescent proteins based on a different family of proteins, which have excitation spectra up to 700nm and emission up to 720nm. Derived from the phytochrome photoreceptor family
found in plants, bacteria, and cyanobacteria, these proteins make it possible to image much deeper into mammalian tissue and with a higher signal to autofluorescence background
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